Staphylococcus aureus and Bacillus anthracis are pathogenic members of the order Bacillales that each represent a considerable threat to global public health. The rise of Staphylococcus aureus strains resistant to all known antimicrobials has the potential to eliminate available treatment options whereas the successful use of B. anthracis as an agent of bioterror threatens national security. Identifying novel therapeutic targets against these organisms is critical to our continued ability to successfully protect agains these infections. Promising antimicrobial targets are bacterial systems involved in stress sensing and detoxification as both processes are required for infection. Alterations in gene expression in response to stress can be controlled by signal transduction systems known as two-component regulatory systems (TCS). Bacteria typically encode many TCSs that are responsible for recognizing and responding to different signals. However, it is not known to what extent the signaling pathways of distinct TCSs cross-regulate to integrate multiple signals from the environment. In addition, the mechanisms by which bacteria maintain signal purity and prevent unwanted cross-regulation are not understood. In this application we describe the identification of cross-regulating TCSs that integrate the bacterial response to heme and cell envelope stress linking bacterial growth in blood to resistance to antimicrobial agents. This discovery represents one of the first examples of TCS cross-regulation that has been described in a bacterial pathogen. In S. aureus and B. anthracis, the avoidance of heme-mediated toxicity is facilitated by the heme- dependent activation of the heme sensor system TCS (HssRS). HssRS senses heme and activates expression of the heme regulated transporter (HrtAB) which is an efflux pump that protects against heme stress. We recently made the exciting discovery that a previously uncharacterized B. anthracis TCS senses cell envelope stress and cross-signals with HssRS; therefore, we have named this system HssRS interfacing TCS (HitRS). HitRS also activates a previously unstudied efflux pump which we have named HitPQ, implicating HitPQ in protection against cell envelope stress. Based on these data, we propose a model whereby (i) HssRS and HitRS sense heme and cell envelope stress to enable a coordinated response to these distinct stressors, (ii) this response results in the efflux of toxic molecules from the cell, and (iii) the integrity of this synchronized stress response is required fr pathogenesis and engenders bacterial pathogens with increased resistance to antimicrobials upon exposure to blood. We will utilize genetics, biochemistry and animal infection experiments to determine the mechanism by which HssRS and HitRS sense stress, define the functional role of HitRS/HssRS crosstalk, and elucidate the mechanism by which HitPQ and HrtAB protect against toxicity. Due to the fundamental requirement for integrating stress sensing and signal transduction by virtually all bacteria, these studies will be universally applicable across the bacterial kingdom.